Immune-mediated gastrointestinal disorders encompass a wide range of debilitating gastrointestinal diseases of various etiologies. One such immune-mediated gastrointestinal disorder, inflammatory bowel disease (IBD), is the collective term used to describe two gastrointestinal disorders of unknown etiology: Crohn's disease (CD) and ulcerative colitis (UC). The course and prognosis of IBD, which occurs world-wide and afflicts millions of people, varies widely. Onset of IBD occurs predominantly in young adulthood, with diarrhea, abdominal pain, and fever being the three most common presenting symptoms. The diarrhea may range from mild to severe and in ulcerative colitis often is accompanied by bleeding. Anemia and weight loss are additional common signs of IBD. Ten to fifteen percent of all patients with IBD will require surgery over a ten year period. In addition, patients with IBD are at increased risk for the development of intestinal cancer. Symptoms such as an increased occurrence of psychological problems, including anxiety and depression, are perhaps not surprising given that IBD is often a debilitating disease that strikes people in the prime of life.
Crohn's disease (CD) is a major form of IBD leading to a chronic granulomatous inflammation that can occur anywhere in the alimentary tract (Podolsky, N. Engl. J. Med. 325:928-937 (1991)). Although the precise pathogenesis is still unknown, recent data suggest that genetic factors (e.g., NOD2 mutations), environmental factors (e.g., bacterial antigens), and an activation of the mucosal immune system play a major pathogenic role (Podolsky, supra; Beutler, Immunity, 15:5-14 (2001); Neurath et al., Nat. Med. 8:567-573 (2002); MacDonald et al., Scand. J. Immunol. 51:2-9 (2000); Shanahan, Lancet 359:62-69 (2002); Hugot et al., Nature 411:599-603 (2001); Ogura et al., Nature 411:603-606 (2001); Targan et al., N. Engl. J. Med. 337:1029-1035 (1997); Elson et al., Gastroenterol. 109:1344-1367 (1995)). In the chronic phase of this disease, cytokines produced by macrophages and T lymphocytes in the lamina propria have been shown to play a key pathogenic role (Atreya et al., Nat. Med. 6:583-588 (2000); Breese et al., Immunology 78:127-131 (1993); Plevy et al., J. Immunol. 159:6276-6282 (1997)). In particular, it has recently been demonstrated that proinflammatory cytokines such as IL-6 and IL-12 may cause T cell resistance against apoptosis in CD that leads to inappropriate lymphocyte accumulation in the gut and disease perpetuation (Atreya et al., supra; Boirivant et al., Gastroenterol. 116:557-565 (1999)).
The azathioprine molecule is composed of two moieties: mercaptopurine and an imidazole derivative (Lennard, Eur. J. Clin. Pharmacol. 43:329-335 (1992); Hoffmann et al., J. Am. Chem. Soc. 123:6404-6409 (2001)). After oral administration and absorption, the pro-drug azathioprine undergoes approximately 90% conversion to 6-MP by non-enzymatic attack by sulfhydryl-containing compounds such as glutathione or cysteine that are present in every mammalian cell (Lennard, supra; Kroplin et al., Eur. J. Clin. Pharmacol. 56:343-345 (2000)). 6-MP is then enzymatically converted by xanthine oxidase to 6-thiouric acid, by thiopurine S-methyltransferase (TPMT) to 6-methyl-MP (6-MMP), and by hypoxanthine phosphoribosyl transferase (HPRT) to 6-thioguanine (6-TG). Whereas the TPMT pathway appears to be important for azathioprine-mediated side effects such as myelosuppression, the 6-TG generated by the HPRT pathway most likely mediates the immunosuppressive properties of 6-MP (Lennard, supra). In particular, lymphocytes have been shown to enzymatically convert 6-MP to 6-TG (Van Os et al., Gut 39:63-68 (1996)).
Azathioprine and its metabolite 6-mercaptopurine (6-MP) are widely used as immunosuppressive and anti-inflammatory agents in organ transplantation and the treatment of chronic inflammatory diseases as well as in the treatment of leukemia. For instance, azathioprine has been therapeutically used in kidney (McGeown et al., Lancet 1:983-991 (1988)) and heart (Andreone et al., J. Heart Transplant. 5:13-19 (1986)) transplantation and various autoimmune and chronic inflammatory diseases, such as multiple sclerosis (Lancet 2:179 183 (1988)), rheumatoid arthritis (DeSilva et al., Ann. Rheum. Dis. 40:560-568 (1981)), systemic lupus erythematosus (Ginzler et al., Arthritis Rheum. 18:27-35 (1975)), primary biliary cirrhosis (Christensen et al., Gastroenterol. 89:1084-1091 (1985)), and IBD (Candy et al., Gut 37:674-678 (1995); Bouhnik et al., Lancet 347:215-219 (1996); D'Haens et al., Gastroenterol. 112:1475-1481 (1997); Lewis et al., Gastroenterol. 118:1018-1024 (2000); Present et al., N. Engl. J. Med. 302:981-987 (1980)). Azathioprine is considered the gold standard of immunosuppressive therapy for CD (Present et al., supra; Sandborn et al., Gastroenterol. 117:527-535 (1999)). Interestingly, it has been shown that patients with CD who were treated with azathioprine for only a few weeks did not respond to therapy, suggesting that this drug requires prolonged periods of time to achieve clinical responses (Pearson et al., Ann. Intern. Med. 123:132-142 (1995); Ewe et al., (1993) Gastroenterol. 105:367-376 (1993)). Furthermore, a recent meta-analysis demonstrated that the odds ratio of response to azathioprine in this disease increased with the cumulative dose administered (Pearson et al., supra).
Although azathioprine has been in clinical use for about four decades (Bean, Med. J. Aust. 2:592-593 (1962)), its precise mechanisms of action are still unknown. For example, inhibition of purine nucleotide biosynthesis with suppression of DNA and RNA synthesis and down-regulation of B and T cell function have been suggested as major therapeutic mechanisms of azathioprine (Dimitriu et al., J. Immunol. 121:2335-2339 (1978); Lennard, supra; Röllinghoff et al., Clin. Exp. Immunol. 15:261-269 (1973); Abdou et al., Clin. Exp. Immunol. 13:55-64 (1973); Bach et al., Clin. Exp. Immunol. 11:89-98 (1972)). However, it is well accepted that a general inhibition of nucleic acid synthesis is not sufficient to explain the specific effects of azathioprine and 6-MP on the immune system (Elion, Ann. New York Acad. Sci. 685:400-407 (1993)). Furthermore, azathioprine and 6-MP require very high dosages to suppress proliferation of primary T cells in vitro that are outside of clinically relevant dosages in IBD patients (Quemeneur et al., J. Immunol. 170:4986-4995 (2003)). Taken together, these data make it very likely that another specific mechanism underlies the immunosuppressive properties of azathioprine.
Since unchecked proliferation of lymphocytes may provoke the risk of developing chronic inflammatory or autoimmune diseases such as IBD, the immune system controls efficient elimination of activated lymphocytes in a process known as apoptosis (Scaffidi et al., Curr. Opin. Immunol. 11:277-285 (1999); Lenardo et al., Annu. Rev. Immunol. 17:221-253 (1999)). This is particularly important for the mucosal immune system, since a resistance of lamina propria cells to apoptosis may lead to chronic inflammatory responses in the gut (Neurath et al., Trends Immunol. 22:21-26 (2001)). Thus, there is a need in the art for novel therapeutic methods of inducing the apoptosis of immune cells such as lymphocytes in patients with IBD. There is also a need in the art for novel methods of monitoring, predicting, and/or optimizing clinical responsiveness to therapy in patients with IBD. The present invention satisfies these needs and provides related advantages as well.